Project Summary The research proposed here seeks to address how, on a molecular level, environmental sensing proteins transmit the sensed signal to the biologically active domain of the protein. The system to be studied is a monomeric, soluble, histidine kinase from the marine bacterium Erythrobacter litoralis HTCC2594 (EL346): specifically, how the protein senses blue light using a sensor LOV (light-oxygen-voltage) domain (a type of PAS [Per-ARNT-Sim] domain) and passes on the signal to a kinase domain. The aim of this research is to identify a more generally applicable mechanism for activation than that previously proposed for dimeric histidine kinases. Specific research goals are (1) to figure out what determines whether a sensor kinase is monomeric or dimeric, (2) to determine the molecular mechanism of activation of the kinase domain, and (3) to outline what types of small molecules different PAS domains bind. First, proteins homologous to EL346 will be found, their oligomeric state determined, and high-resolution structures of monomeric and dimeric proteins compared. Second, high-resolution structures will be determined for active and inactive histidine kinases and compared. Thirds, a fragment library of small molecules will be screened against a representative set of PAS domains to tease out what types of PAS domains bind what types of ligands. The research training plan includes working under Dr. Kevin Gardner's supervision to gain expertise in integrating different structural biology techniques with biochemical data to fully describe a biological system on a molecular scale. Sensor histidine kinases are part of two-component systems used by bacteria, plants, and fungi to sense and respond to different cues in the environment. Because humans lack these two-component systems, these bacterial systems are attractive targets for targeted antibacterial drugs. However, high-resolution structural information and a mechanistic understanding of signal transmission are required to design effective therapeutics. The research proposed here will provide this understanding. Other sensor kinases use the related PAS domain as a sensor for small molecules; these systems are attractive as starting points for biosensors (cellular, diagnostic, or environmental). The expected findings on PAS domain compound binding can be used to design such biosensors. Thus, the outcomes of the proposed experiments will not only increase researchers' understanding of signal transduction systems in general, but also provide a foundation for the development of new biosensors and therapeutics and impact how diseases are treated.